A standard antibody is a tetrameric structure consisting of two identical immunoglobulin heavy chains and two identical light chains. The heavy and light chains of an antibody consist of different domains. Each light chain has one variable domain and one constant domain, while each heavy chain has one variable domain and three or four constant domains. Alzari, P. N., Lascombe, M.-B. & Poljak, R. J. (1988). Three-dimensional structure of antibodies. Ann. Rev. Immunol. 6, 555-580. Each domain consists of about 110 amino acid residues. Each domain is also folded into a characteristic β-sandwich structure formed from two β-sheets packed against each other (the immunoglobulin fold). The variable heavy and variable light domains each have three complementarity determining regions (CDR1-3) that connect the β-strands at one end of the domains. The variable regions of both the light and heavy chains generally contribute to antigen specificity, although the contribution of the individual chains to specificity is not always equal. Hence, antibody molecules are large and complex.
Antibody molecules have evolved to bind to a large number of molecules by using six randomized loops (CDRs). However, the size and the existence of six different loops on separate polypeptides constitute a hurdle to molecular manipulations that might otherwise be used to improve the structure, stability and binding properties of antibodies. Moreover, while antibodies are widely used in medical research, industrial processes and in diagnostics, they are expensive and difficult to obtain. They also lack suitable stability for long shelf life.
What would be useful is a smaller, more stable binding agent that can easily be manipulated by standard cloning procedures and that could be produced in cultured host cells, rather than in animals. Such new types of binding agents would ideally have the positive features of antibodies (e.g., high specificity and affinity for binding a distinct target) but few of the negative aspects of antibodies (e.g., instability and difficulty of production). Moreover, new procedures are also needed for large-scale preparation of such binding agents in cultured cells that would avoid the time and expense of using animals.